The principal aims of this research are to elucidate the ionic mechanisms by which black widow spider venom (BWSV) and its purified protein, Alpha-latrotoxin: 1) stimulate the release of quanta of acetylcholine (ACh), and 2) deplete nerve terminals of their synaptic vesicles. The hypothesis to be tested is that BWSV acts by inducing changes in the permeability of the nerve terminal axolemma to monovalent and divalent cations. Two experimental approahces will be made to this study of permeability change. The first uses micropipettes to implae presynaptic terminals of the avian ciliary ganglion to measure directly the changes in resting potential, action potential, and membrane resistance that occur during the action of BWSV. This method allows a direct test of the hypothesis, for example, that the toxin acts on the channels through which Ca++ enters a nerve terminal thereby triggering the release of ACh. The second approach examines the effects of BWSV at the frog neuromuscular junction, focusing on the observation that the depletion of synaptic vesicles caused by low doses of BWSV is most pronounced when the extracellular concentration of Ca++ is very low. This result has suggested that endocytosis requires Ca++. Newly developed techniques of fluctuation analysis will be applied to the stochatic variations in electrical potential recorded at vigorously secreting junctions. Quantitative estimates will be obtained of the rates of quantal secretion under different ionic conditions, some of which lead to vesicle depletion, some of which do not. Analysis of the rates of secretion as functions of time under these different conditions will provide insights into the kinetics of both the endocytosis of synaptic vesticles and the refilling of the newly formed vesicles with ACh. Together, these two approaches will reveal the specific effects of BWSV on nerve terminal permeability, and provide information about the role of Ca++ and other factors in the release of ACh and the recycling of synaptic vesicles.